Preparation of a Convertible Spacer Containing a Disulfide Group for Versatile Functionalization of Oligonucleotides
Bibliographic record
Abstract
Abstract The protocols described in this article provide details regarding the synthesis and characterization of a disulfide containing linker phosphoramidite for terminal functionalization of synthetic oligonucleotides. The linker is first synthesized from 6‐mercaptohexanol in two steps and is incorporated at the 5′ end of short DNA oligonucleotides using automated solid‐phase synthesis. The linker contains a terminal tosylate group which is post‐synthetically displaced by altering the deprotection conditions to yield a variety of functional handles (N3, NH2, OMe, SH) or alternatively, the tosylate can be displaced directly with primary amines such as tert‐butylamine. The linker system is also compatible with RNA oligonucleotides enabling the introduction of various functional handles (N3, NH2). The protocol outlined in this procedure provides access to a versatile linker for the terminal functionalization of oligonucleotides containing a disulfide bond which may serve useful in the synthesis of reduction‐responsive oligonucleotide conjugates. As a proof of concept, in this protocol the linker is used to modify a dT10 oligonucleotide and then conjugated by copper(I)‐mediated azide‐alkyne cycloaddition (CuAAC) to an alkyne‐modified poly(ethylene glycol) which shows concentration dependent release of the oligonucleotide upon treatment with 1,4‐dithiothreitol, a reducing agent. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Preparation of disulfide linker phosphoramidite 3 Basic Protocol 2: Synthesis, functionalization, and characterization of DNA oligonucleotides containing disulfide linker phosphoramidite 3 Basic Protocol 3: Displacement of terminal tosylate functionalized DNA with primary aliphatic amines Basic Protocol 4: Synthesis of oligonucleotide‐PEG conjugate Support Protocol: Preparation of PEG‐alkyne
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.003 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".